Preparation of polyvinyl pyrrolidone stationary phase and its application in separation and analysis of flavonoids

Abstract Polyvinylpyrrolidone (PVP) is known as a polymer with special adsorption properties for polyphenolic compounds. In this paper, chemical‐bonded stationary phase (PVP@Silica) was prepared and applied to the separation of flavonoids. System constants determined by linear solvation energy relationships with methanol–water mixtures as the mobile show that the column can provide stronger hydrogen bonding and π–π interaction than C18 column. Under reversed‐phase conditions, traditional flavonoids have much longer retention on PVP@Silica than on C18 column with methanol–water mobile phase due to multimodal retention mechanism, including hydrogen bonding and π–π interaction. In addition, a U‐shaped retention curve was observed in acetonitrile–water mobile phase because of the enhanced hydrogen bonding under the high proportion of acetonitrile. The selectivity to polyhydroxy structure gives the stationary phase unique separation ability for flavonoids. The separation orthogonality was further investigated by a sample set containing 33 flavonoids with different substitution structures. The (RP‐PVP@Silica)‐(RP‐C18) system exhibited 60% orthogonality metric (OM) for these flavonoids with methanol–water mobile phase. A high OM of 63.5% was achieved in (RP‐PVP@Silica)‐(HILIC‐PVP@Silica) system. Finally, PVP@Silica was applied to the purification of total flavonoids in Ginkgo biloba extract (GBE). The offline (RP‐PVP@Silica)‐(RP‐C18) two‐dimensional separation system was used for the analysis of flavonoids in GBE.